This invention relates to the treatment of pathological conditions and diseases involving abnormal cellular proliferations, such as tumors, cancers, restenosis, hyperplasias, corneal haze and cataracts by providing mutated cyclin G1 protein to an affected animal, thereby inhibiting the function of native cyclin G1 protein. More particularly, this invention relates to the treatment of such conditions and diseases by administering to the animal an expression vehicle, such as a retroviral vector or an adenoviral vector, which comprises a polynucleotide encoding mutated cyclin G1 protein.
Genes encoding a new class of proteins known as cyclins have been identified as a new class of protooncogenes, and cyclin-dependent kinase (or Cdk) inhibitors have been identified as tumor suppressors, thereby uniting the molecular mechanisms of cellular transformation and tumorigenesis with the enzymology governing cell cycle control. (Hall, et al., Curr. Opin. Cell Biol., Vol. 3, pgs. 176-184 (1991); Hunter, et al., Cell, Vol. 79; pgs. 573-582 (1994); Elledge, et al., Curr. Opin. Cell Biol., Vol 6, pgs. 874-878 (1994); Peter, et al., Cell, Vol. 79, pgs. 181-184 (1994)). The sequential expression of specific cyclins and the essential functions of specific Cdk complexes have been defined (Wu, et al., Int. J. Oncol., Vol. 3, pgs. 859-867 (1993); Pines, et al., New Biologist, Vol. 2, pgs. 389-401 (1990); Pines, Cell Growth and Differentiation, Vol. 2, pgs. 305-310 (1991); Reed, Ann. Rev. Cell Biol., Vol. 8, pgs. 529-561 (1992); Sherr, Cell, Vol. 79, pgs. 551-555 (1994)), thereby providing direct links to the fundamental mechanisms of DNA replication, transcription, repair, genetic instability, and apoptosis. (D""Urso, et al., Science, Vol. 250, pgs. 786-791 (1990); Wu, et al., Oncogene, Vol. 9, pgs. 2089-2096 (1994); Roy, Cell, Vol. 79, pgs. 1093-1101 (1994); Meikrantz, et al., Proc. Nat. Acad. Sci., Vol. 91, pgs. 3754-3758 (1994)).
Metastatic carcinoma is an important target for gene therapy as the disease is associated with poor outcome. Colorectal cancer, for example, is the second leading cause of cancer death in the United States after lung cancer, followed by breast and pancreatic cancer (Silberberg et al., Cancer Clin., Vol. 40, pgs. 9-26 (1990)). Of these carcinomas, pancreatic cancer has the worst prognosis. The median survival of patients with metastatic pancreatic cancer is three to six months and virtually all the patients are dead within a year (Merrick et al., Gastrenterol. Clin. N. Amer., Vol. 19, pgs. 935-962 (1990)). Approximately 40% of patients will have metastatic disease either to the liver or the peritoneal cavity or both at the time of diagnosis. Chemotherapy for metastatic disease is ineffective despite multimodal therapy. Hence, alternative approaches to metastatic carcinoma would be desirable.
Wu, et al., (Oncol. Reports, Vol. 1, pgs. 705-711 (1994)), discloses the deduced amino acid sequence and cDNA sequence for human cyclin G1 protein. Wu, et al., also disclose that higher levels of cyclin G1 expression were found in osteosarcoma cells and in Ewing""s sarcoma cells than in normal diploid human fibroblasts. Although Wu, et al., state that the overexpression of cyclin G1 protein in human osteosarcoma cells provides a potential link to cancer, Wu, et al., do not disclose the treatment of cancer by interfering with or inhibiting the function of cyclin G1 protein in cancer cells.
Atherosclerosis, a principal cause of both myocardial and cerebral infarction, is responsible for xcx9c50% of all mortality in the United States and Europe (Ross, Nature, Vol. 362. pgs. 801-809 (1993); Murray and Lopez, The Global Burden of Disease, Harvard University Press, Cambridge, Mass. (1996)). In addition to bypass grafting and endarterectomy, percutaneous transluminal coronary angioplasty (PTCA) has become standard treatment for vascular stenosis (Fitz Gibbon, et al., Can. J. Cardiol., Vol. 12, pgs. 893-900 (1996)). While the success rate of the initial PTCA has increased to well over 90%, the long-term efficacy of the procedure is limited by the eventual development of neointimal hyperplasia and restenosis in xcx9c30-50% of patients (Glagov, Circulation, Vol. 89 pgs. 2888-2891 (1994); Schwartz et al., Am. Coll. Cardiol., Vol. 17, pgs. 1284-1293 (1992); Myers et al., Wound Healing Responses in Cardiovascular Disease, Weber, ed, Futura Publishing Co., Mt. Kisco, N.Y., pgs. 137-150 (1995); Chen, et al., J. Clin, Invest., Vol 99, pgs. 2334-2341 (1997)), often to such an extent that a second PTCA is necessary (Kirchengast, et al., Cardiovasc. Res., Vol. 39, pgs. 550-555 (1998)). To date, no pharmacological strategy has been sufficiently effective to warrant its widespread use (Herrman et al., Drugs, Vol. 46, pgs. 18-52 (1993); De Meyer and Bult, Vascul. Med. Vol. 2, pgs. 179-189 (1997)). Thus, the control of neointima formation represents a major goal of contemporary research in vascular biology (Schwartz, et al., The Intima. Circulation Res., Vol. 77, pgs. 445-465 (1995)) and a model system for the development of new molecular medicines (Gibbons, et al., Science, Vol. 272, pgs. 689-693 (1996)).
The high degree of complexity and redundancy in growth factor signaling pathways has prompted the examination of conserved cell cycle control pathways in the design of novel cytostatic therapies (Barr and Leiden, Trends Cardiovasc. Med., Vol 4 pg. 57 (1994); Andres, Int. J. Molecular. Med., Vol. 2, pgs. 81-84 (1998); Braun-Dullaeus et al., Circulation, Vol. 98, pgs. 82-89 (1998)). Consequently, a number of novel gene therapy approaches to inhibit SMC proliferation and neointima formation have focused on specific cell cycle control elements, including oligodeoxynucleotides representing antisense constructs of cyclin-dependent protein kinase (CDK) subunits (Morishita et al., Proc. Nat. Acad. Sci., Vol. 90, pgs. 8474-8478 (1993), Morishita, et al. J. Clin. Invest., Vol. 93. pgs. 1458-1464 (1994); Abe, 1994), adenoviral vectors bearing Cdk inhibitors (Chang et al., Science, Vol. 267, pgs. 518-522 (1995); Chen et al., 1997;) or vectors bearing constitutively active forms of the Rb protein (Chang et al., J. Clin. Invest., Vol. 96 pgs , 2260-2268 (1995); Smith et al., Exp. Cell Res. Vol. 230, pgs. 61-68 (1997)). Other studies have employed molecular xe2x80x9cdecoyxe2x80x9d oligodeoxynucleotide strategies directed against the transcription factor E2F (Morishita, et al., Proc. Nat. Acad. Sci., Vol. 92, pgs. 5855-5859 (1995)), which regulates the induction of multiple cell cycle control genes. The reported efficacy of these experimental approaches supports the concept that, cell cycle control elements which are selectively up-regulated in neointima lesions would represent strategic therapeutic targets.
Recent studies have characterized the up-regulation of cyclin G1, an inducible cell cycle control element (Tamura et al., Oncogene, Vol. 8, pgs. 2113-2118 (1993); Wu, et al., 1994; Home, et al., J. Biol. Chem., Vol. 271, pgs. 6050-6061 (1996); Morishita et al., 1995), following balloon catheter injury in rodents (Zhu et al., 2000; submitted) and non-human primates (Kaijin Wu et al., 1999; submitted). Enforced expression of cyclin G1 in transfected cells in vitro accelerates the cell cycle and promotes clonal expansion (Smith et al, Exp. Cell, Res. Vol. 230, pgs. 61-68 (1997),) while blockade of cyclin G1 expression by antisense strategies induces cytostasis and cytolysis (Skotzko, et al, Cancer Res., Vol. 55 pgs. 61-68 (1995); Chen, et al., Hum. Gene Ther., Vol. 8, pgs. 1667-1674 (1997); Hung, et al., Int. J. Pediatr. Hematol. Oncol., Vol. 4, pgs. 317-325 (1997).) In the context of SMC proliferation, it has been shown (i) that an antisense cyclin G1 retroviral vector concentrated to sufficiently high titer (108 cfu/ml) inhibited the survival and proliferation of transduced rat (Zhu, et al., Circulation, Vol. 46 pgs., 628-635 (1997)) and primate vascular SMCs (unpublished observations), and (ii) that intraluminal delivery of this concentrated antisense cyclin G1 vector in balloon-injured rat carotid arteries produced a significant reduction in neointima formation in vivo (Zhu, et al., 1997). Therefore, cyclin G1 appears to be both a pertinent and advantageous locus for therapeutic intervention in the management of vascular restenosis.
Applicants have discovered that by interfering with and/or inhibiting the function of cyclin G1 protein in cancer cells, one may inhibit, prevent, or destroy the growth and/or survival of such cancer cells. Thus, the present invention is directed to the treatment of pathological conditions and diseases involving abnormal cellular proliferations by inhibiting the function of cyclin G1 protein, through the administration of mutant cyclin G1 proteins to such proliferating cells in an animal. Such pathological conditions and diseases include, but are not limited to, cancers, tumors, hyperplasias, restenosis, corneal haze, and cataracts. In preferred embodiments, the mutant or variant cyclin G1 protein is administered to the affected animal by delivery of expression vehicles comprising polynucleotides encoding mutated cyclin G1 proteins. Such expression vehicles include, but are not limited to viral vectors such as retroviral vectors and adenoviral vectors, and synthetic vectors. Animals that can be treated beneficially by the methods of the invention include, but are not limited to, mammals, including human and non-human primates, dogs, cats, horses, cattle, and sheep.